In the cosmetics field, microbial activity can lead to undesired changes, such as body odor, dandruff formation or open inflammations and spots. The prior art discloses the use of antimicrobial and/or antifungal substances in preparations, e.g. in cosmetics or in medicine. However, the development of new antimicrobial and/or antifungal compositions is constantly necessary due to the development of bacterial or fungal strains that are resistant to active ingredients. Of particular interest is the development of novel antimicrobial and anti-fungal compounds, respectively, which do not show negative effects on the human health and which show a good activity spectrum against harmful germs.
Diverse antimicrobial peptides are already described in the literature and summarized in reviews (Hancock, R. E. W. and Lehrer, R. 1998 in Trends in Biotechnology, 16: 82-88; Hancock, R. E. W. and Sahl, H. G. 2006 in Nature Biotechnology, 24: 1551-1557).
Fusion peptides, which combine two effective peptides, are likewise described in the literature. Wade et al. report on the antibacterial effect of various fusions of cecropin A from Hyalophora cecropia and bee toxin melittin (Wade, D. et al., 1992, International Journal of Peptide and Protein Research, 40: 429-436). Shin et al. describe the antibacterial effect of a fusion peptide of cecropin A from Hyalophora cecropia and magainin 2 from Xenopus laevis, consisting of 20 amino acids. Cecropin A consists of 37 amino acids and exhibits activity toward Gram-negative bacteria, but lower activity toward Gram-positive bacteria. Magainin 2 consists of 23 amino acids and is active against bacteria but also tumor cell lines. Compared to the fusion of cecropin A and melittin, this fusion exhibits considerably lower hemolytic activity for comparable antibacterial effect (Shin, S. Y. Kang, J. H., Lee, M. K., Kim, S. Y., Kim, Y., Hahm, K. S., 1998, Biochemistry and Molecular Biology International, 44: 1119-1126).
US 2003/0096745 A1 and U.S. Pat. No. 6,800,727 B2 claim these fusion peptides consisting of 20 amino acids and variants of this fusion which, as a result of the exchange of amino acids, in particular of positively charged amino acids and hydrophobic amino acids, are more positively charged and more hydrophobic.
Further developments of this cecropin-A-magainin-2 fusion peptide are described by Shin et al. 1999. It is found here that the construct P18 (HT2, SEQ ID NO:3) had lower hemolytic activity compared to the starting fusion, although the antibacterial activity toward Escherichia coli and Bacillus subtilis was not impaired (Shin et al. 1999 Journal of Peptide Research, 53: 82-90).
Furthermore, Shin et al. 2001 investigated the activity of the construct P18 and analogous constructs for their antibacterial activity also on other target bacteria such as Pseudomonas aeruginosa or Proteus vulgaris (Shin et al., 2001, Journal of Peptide Research, 58: 504-514).
First experiments on the effectiveness of P18 against fungi was published in January 2002 by Shin et al. Here, better effectiveness of the peptide P18 towards Candida albicans compared to magainin 2 was established (Shin et al., 2002, Biochemical and Biophysical Research Communications, 290: 558-562).
In addition to Candida albicans, the Ascomycetes Aspergillus flavus, Fusarium oxysporum and the Basidiomycete Trichosporon beigelii were inhibited with P18 and variants of the peptide. However, the most effective inhibition was achieved with P18 (Lee et al., 2004, Biotechnology Letters, 26: 337-341). An inhibition of lipophilic fungi, in particular of the genus Malassezia is not taught. Furthermore, no experiments are described which demonstrate a more effective effect of cecropin-magainin fusions compared to known, commercial antifungal substances.
The genus Malassezia are lipophilic fungi which in part belong to the normal resident flora on human skin. Many representatives are obligatorily lipophilic, only M. pachydermatis is described as optionally lipophilic. Nevertheless disorders which are associated with Malassezia ssp. are described. These include pityriasis versicolor, atopic dermatitis, pityriasis capitis, seborrhea and pityrosporum folliculitis or malassezia folliculitis, for which Malassezia ssp. are pathogens. (Cutsem et al., 1990, Journal of the American Academy of Dermatology, 22: 993-998, Nenoff et al., 2007, Aktuelle Dermatologie, 33: 26-32). For treatment, antimycotic, fungicidal agents are usually used. Examples thereof are ketokonazole, climbazole, zinc pyrithiones, piroctone olamine, selenium sulfides or some natural extracts (e.g. like juniper oil, rosemary oil), which are often added in the single-digit %, (w/w) range, for example, in antidandruff shampoo formulations.
However, some of the described substances for inhibiting, treating or preventing dandruff are either toxicologically unacceptable, or the cosmetic preparations are not effective enough (Kosmetische Medizin 5+6/2006).
The species Malassezia furfur is less sensitive toward several antimycotics compared to other Malassezia species (Nenoff et al., 2007, Aktuelle Dermatologie, 33: 26-32). It is likewise described that substances which considerably inhibit the growth of Candida albicans even at low concentrations, only prevent the growth of Malassezia furfur at high concentrations, if at all. Thus, Nenoff et al. describe the inhibition of Candida albicans by phytosphingosines at concentrations of about 152-269 μg/ml whereas an inhibition of M. furfur only took place at approximately 25-fold higher concentrations (6250 μg/ml) for basic phytosphingosines, or M. furfur appeared to be resistant to phytosphingosine salts (Nenoff et al., 2002, Acta Derm Venerol, 82:170-173).
Similar observations were made for the peptide cilofungin by Hanson et al. 1989. While 76% of the C. albicans strains did not grow at a cilofungin concentration of 0.62 μg/ml, the obligatorily lipophilic M. furfur was not inhibited at these concentrations. The minimum inhibitory concentration of cilofungin for M. furfur was not stated. It was evidently not within the investigated range between 0.31 μg/ml and 40 μg/ml (Hanson et al., 1989, Antimicrobial agents and chemotherapy, 33:1391-1392).
In 2006, Lopez-Garcia et al. describe the growth inhibition of M. furfur with Cecropin P1 and magainin 2. Here, a moderate antifungal effect of cecropin P1 and a comparably better effect of magainin 2 at a concentration of 25 μM were found (Lopez-Garcia et al., 2006, Journal of Antimicrobial Chemotherapy, 57:877-882).
EP-A-0 866 804 describes the use of beta-defensins from human skin as active ingredient in cosmetic or pharmaceutical formulations. However, an internal comparison with antifungal active ingredients of the prior art is not made.
WO-A-00/032220 describes the use of a fungal polypeptide as antifungal active ingredient for the treatment of dandruff. However, an internal comparison with antifungal active ingredients of the prior art is not made here either.
US 2003/0096745 describes a polypeptide of the sequence KWKKLLKKPPPLLKKLLKKL (SEQ ID NO: 4739) with antibacterial and antifungal activity toward certain microorganisms. Antifungal activity was shown toward Candida albicans and Tricosphoron beigelii. Cosmetic applications, in particular for the treatment of dandruff, are not proposed.
However, the use of antimicrobial substances, particularly regular use, can lead to intolerance in people, or even to health damage. Intolerances may be skin reddening, irritations or sensitizations. Systemic absorption into the human body can lead to impairment of body functions. Particularly regular use of some antimicrobial substances can lead to an increase in their concentration. One known example is parabens (Dabre et al., 2004, Journal of Applied Toxicology, 24: 5-13). Depending on the application, an accumulation in the human body or in the environment may thus result.
In addition, excessive and inappropriate use of antimicrobial substances result time and again in resistance of the target organisms.
There is therefore a need to provide novel cosmetic antimicrobial compositions which firstly have antifungal effectiveness and are suitable for avoiding or treating dandruff. In particular, these should not accumulate in the body since they can be degraded in the natural surroundings.
There is in particular a need to provide antifungal active ingredients which are more effective against dandruff than the customary antidandruff agents known hitherto and in particular are effective against the dandruff fungus Malassezia furfur and other Malassezia ssp. Preferably, the natural skin flora should not be impaired, and the active ingredients should also not accumulate in the body or in the environment, but be degraded in the natural surroundings.
It was therefore an object of the present invention to provide a novel, effective but biodegradable active ingredient for avoiding, inhibiting and/or treating dandruff, in particular scurf. Advantageously, active ingredient compounds are to be identified which are suitable for producing cosmetic and/or dermatocosmetic formulations or preparations. Furthermore, systemic absorption of the active ingredient should be avoided. Additionally, it should be ensured that the preparations have low or no cytoxicity. In particular, the active ingredient should be effective against the yeast fungus Malassezia furfur, and also against other Malassezia ssp., in particular, obligatorily lipophilic species.